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Pancreatic cancer is associated with a pronounced fibrotic reaction that was recently shown to limit delivery of chemotherapy. To identify potential therapeutic targets to overcome this fibrosis, we examined the interplay between fibrosis and the key proteinase membrane type 1-matrix metalloproteinase (MT1-MMP, MMP-14), which is required for growth and invasion in the collagen-rich microenvironment. In this article, we show that compared with control mice (Kras+/MT1-MMP−) that express an activating KrasG12D mutation necessary for pancreatic cancer development, littermate mice that express both MT1-MMP and KrasG12D (Kras+/MT1-MMP+) developed a greater number of large, dysplastic mucin-containing papillary lesions. These lesions were associated with a significant amount of surrounding fibrosis, increased α-smooth muscle actin (+) cells in the stroma, indicative of activated myofibroblasts, and increased Smad2 phosphorylation. To further understand how MT1-MMP promotes fibrosis, we established an in vitro model to examine the effect of expressing MT1-MMP in pancreatic ductal adenocarcinoma (PDAC) cells on stellate cell collagen deposition. Conditioned media from MT1-MMP–expressing PDAC cells grown in three-dimensional collagen enhanced Smad2 nuclear translocation, promoted Smad2 phosphorylation, and increased collagen production by stellate cells. Inhibiting the activity or expression of the TGF-β type I receptor in stellate cells attenuated MT1-MMP conditioned medium–induced collagen expression by stellate cells. In addition, a function-blocking anti–TGF-β antibody also inhibited MT1-MMP conditioned medium–induced collagen expression in stellate cells. Overall, we show that the bona fide collagenase MT1-MMP paradoxically contributes to fibrosis by increasing TGF-β signaling and that targeting MT1-MMP may thus help to mitigate fibrosis. Mol Cancer Res; 9(10); 1294–304. ©2011 AACR.

Pediatric intravenous catheter insertion can be difficult in the operating room due to the technical challenges of small diameter vessels and the need to rapidly gain intravenous access in anesthetized children. Few studies have examined factors associated with difficult vascular access in the operating room, especially accounting for the increased possibility to use ultrasound guidance.The primary aim of the study was to identify factors associated with pediatric difficult vascular access in the operating room. Our primary hypothesis was that Black race, Hispanic ethnicity, and ultrasound use would be associated with pediatric difficult vascular access.We performed a retrospective analysis of prospectively collected data from a cohort of pediatric patients who had intravenous catheters inserted in the operating room at an academic tertiary care children's hospital from March 2020 to February 2021. We measured associations among patients who were labeled as having difficult vascular access (2 attempts at access) with demographic, clinical, and hospital factors.12 728 intravenous catheter insertions were analyzed. Multivariable analysis showed significantly higher odds of difficult vascular access with Black non-Hispanic race (1.43, 95% CI: 1.06-1.93, p = .018), younger age (0.93, 95% CI: 0.89-0.98, p = .005), overweight (1.41, 95% CI: 1.04-1.90, p = .025) and obese body mass index (1.56, 95% 95% CI: 1.12-2.17, p = .008), and American Society of Anesthesiologists physical status III (1.54, 95% CI:1.11-2.13, p = .01). The attending anesthesiologist compared to all other practitioners (certified registered nurse anesthetist: (0.41, 95% CI: 0.31-0.56, p .001, registered nurse: 0.25, 95% CI: 0.13-0.48, p .001, trainee: 0.21, 95% CI: 0.17-0.28, p-value.001 with attending as reference variable) and ultrasound use (2.61, 95% CI: 1.85-3.69, p .001) were associated with successful intravenous catheter placement.Black non-Hispanic race/ethnicity, younger age, obese/overweight body mass index, American Society of Anesthesiologists physical status III, and ultrasound were all associated with pediatric difficult vascular access in the operating room.

PDF file - 153K, LY294002 attenuates mast cell mediated CD11b+ cell migration. Representative images of migration of CD11b cells in response to (A) Stempro +SCF (control), (B) LAD-2 CM (LAD-2 conditioned medium), (C) LAD-2 CM + 10 �M LY294002 (LAD-2 CM with 10 �M LY294002) and (D) CM obtained after pretreatment of LAD-2 cells with 10 �M LY294002. (E) Purity of CD11b+ cells obtained after using either biotinylated Rat ant-human CD11b or biotinylated Rat IgG2bK. Black arrows indicate migrated CD11b+ cells. Black scale bar = 20 �m.

PDF file - 173K, Tumor/Margin frequency of CD4 T cells is inversely related to Macrophages and Mast cells. (A) Graphical Representation and (B) Representative images of the CD11b, Mac387, Tryptase, CAE and CD4 cells in tumor and tumor margin area.

PDF file - 54K, Toluidine Blue staining protocol.

PDF file - 55K, Mast cells promote in vitro and in vivo Gr1+ neutrophil and Foxp3+ Tregs migration in PI3K dependent manner. (A) GMMCs promote Gr1+ neutrophil in vitro migration and pretreatment of GMMCs with LY294002 attenuates migration of Gr1+ cells (B) LY294002 in vivo treatment attenuates migration of Gr1+ neutrophils (C) LY294002 in vivo treatment attenuates migration of Foxp3+ Tregs, * P < 0.05 represents the result of ANOVA.

Purpose: To understand signaling pathways that shape inflamed tissue and predispose to cancer is critical for effective prevention and therapy for chronic inflammatory diseases. We have explored phosphoinositide 3-kinase (PI3K) activity in human inflammatory bowel diseases and mouse colitis models.Experimental Design: We conducted immunostaining of phosphorylated AKT (pAKT) and unbiased high-throughput image acquisition and quantitative analysis of samples of noninflamed normal colon, colitis, dysplasia, and colorectal cancer. Mechanistic insights were gained from ex vivo studies of cell interactions, the piroxicam/IL-10−/− mouse model of progressive colitis, and use of the PI3K inhibitor LY294002.Results: Progressive increase in densities of pAKT-positive tumor-associated macrophages (TAM) and increase in densities of mast cells in the colonic submucosa were noted with colitis and progression to dysplasia and cancer. Mast cells recruited macrophages in ex vivo migration assays, and both mast cells and TAMs promoted invasion of cancer cells. Pretreatment of mast cells with LY294002 blocked recruitment of TAMs. LY294002 inhibited mast cell and TAM-mediated tumor invasion, and in mice, blocked stromal PI3K, colitis, and cancer.Conclusion: The PI3K/AKT pathway is active in cells infiltrating inflamed human colon tissue. This pathway sustains the recruitment of inflammatory cells through a positive feedback loop. The PI3K/AKT pathway is essential for tumor invasion and the malignant features of the piroxicam/IL-10−/− mouse model. LY294002 targets the PI3K pathway and hinders progressive colitis. These findings indicate that colitis and progression to cancer are dependent on stromal PI3K and sensitive to treatment with LY294002. Clin Cancer Res; 19(9); 2342–54. ©2013 AACR.

PDF file - 97K, Multiplex ELISA data for human inflammatory chemoattractants.

Purpose: To assess the clinical and pathologic significance of mast cell infiltration in human pancreatic cancer and evaluate crosstalk between mast cells and cancer cells in vitro.Experimental Design: Immunohistochemistry for tryptase was done on 53 pancreatic cancer specimens. Mast cell counts were correlated with clinical variables and survival. Serum tryptase activity from patients with cancer was compared with patients with benign pancreatic disease. In vitro, the effect of pancreatic cancer–conditioned medium on mast cell migration was assessed. The effect of conditioned medium from the human mast cell line, LAD-2, on cancer and normal ductal cell proliferation was assessed by thymidine incorporation. Matrigel invasion assays were used to evaluate the effect of mast cell–conditioned medium on cancer cell invasion in the presence and absence of a matrix metalloproteinase inhibitor, GM6001.Results: Mast cell infiltration was significantly increased in pancreatic cancer compared with normal pancreatic tissue (11.4 ± 6.7 versus 2.0 ± 1.4, P < 0.001). Increased infiltrating mast cells correlated with higher grade tumors (P < 0.0001) and worse survival. Patients with pancreatic cancer had elevated serum tryptase activity (P < 0.05). In vitro, AsPC1 and PANC-1 cells induced mast cell migration. Mast cell–conditioned medium induced pancreatic cancer cell migration, proliferation, and invasion but had no effect on normal ductal cells. Furthermore, the effect of mast cells on cancer cell invasion was, in large part, matrix metalloproteinase–dependent.Conclusions: Tumor-infiltrating mast cells are associated with worse prognosis in pancreatic cancer. In vitro, the interaction between mast cells and pancreatic cancer cells promotes tumor growth and invasion. Clin Cancer Res; 16(8); 2257–65. ©2010 AACR.

PDF file - 125K, (A) Representative images of HT-29 invasion study in response to Stempro + SCF (internal control for LAD-2 CM), LAD-2 CM, CM + 10 �M LY294002 (LAD-2 CM with 10 �M LY294002) and 10 �M LY pretreated CM (CM obtained after treatment of LAD-2 cells with 10 �M LY294002). (B) Representative images of CFSE labeled HT-29 invasion study in response to tumor infiltrating leukocytes (TILs) isolated from UC associated cancer patients. Red arrows show invaded HT-29. Red scale bar = 50 �m, * P < 0.05 represents the result of Student t test.

PDF file - 40K, Multiplex ELISA data for human inflammatory chemoattractants.

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Supplementary Data from Crosstalk between Mast Cells and Pancreatic Cancer Cells Contributes to Pancreatic Tumor Progression

PDF file - 61K, Table.1 Patient Cohorts. Table 2. Ulcerative Colitis Patients Absent Colonic Dysplasia. 3.1.3 Ulcerative Colitis patients with invasive cancer used in this study. Table 4. Patients diagnosed with ulcerative colitis and associated colorectal cancers.

Supplementary Figure 4 from Mast Cell 5-Lipoxygenase Activity Promotes Intestinal Polyposis in APCΔ468 Mice

Supplementary Table 2 from Tgfbr1 Haploinsufficiency Inhibits the Development of Murine Mutant Kras-Induced Pancreatic Precancer

Supplementary Table 1 from Tgfbr1 Haploinsufficiency Inhibits the Development of Murine Mutant Kras-Induced Pancreatic Precancer

Supplementary Figure 1 from Mast Cell 5-Lipoxygenase Activity Promotes Intestinal Polyposis in APCΔ468 Mice

Supplementary Figure 2 from Mast Cell 5-Lipoxygenase Activity Promotes Intestinal Polyposis in APCΔ468 Mice

To dissect the role of constitutively altered Tgfbr1 signaling in pancreatic cancer development, we crossed Elastase-KrasG12D (EL-Kras) mice with Tgfbr1 haploinsufficient mice to generate EL-Kras/Tgfbr1+/− mice. Mice were euthanized at 6 to 9 months to compare the incidence, frequency, and size of precancerous lesions in the pancreas. Only 50% of all EL-Kras/Tgfbr1+/− mice developed preinvasive lesions compared with 100% of EL-Kras (wild-type Tgfbr1) mice. The frequency of precancerous lesions was 4-fold lower in haploinsufficient than in control mice. Paradoxically, the precancerous lesions of EL-Kras/Tgfbr1+/− mice were considerably larger than those in EL-Kras mice. Yet, the mitotic index of precancerous cells and the observable levels of fibrosis, lipoatrophy, and lymphocytic infiltration were reduced in EL-Kras/Tgfbr1+/− mice. We conclude that Tgfbr1 signaling promotes the development of precancerous lesions in mice. These findings suggest that individuals with constitutively decreased TGFBR1 expression may have a decreased risk of pancreatic cancer. [Cancer Res 2009;69(24):9169–74]

Supplementary Table Legends 1-2 from Tgfbr1 Haploinsufficiency Inhibits the Development of Murine Mutant Kras-Induced Pancreatic Precancer

Supplementary Figure 3 from Mast Cell 5-Lipoxygenase Activity Promotes Intestinal Polyposis in APCΔ468 Mice

The Pediatric Anesthesia COVID-19 Collaborative (PEACOC) is a research network to advance the care of children during the pandemic. Here we calculate the prevalence of coronavirus disease 2019 (COVID-19) among children undergoing anesthesia, look at prevalence in the population data from the Centers for Disease Control and Prevention (CDC), and assess independent risk factors for infection.This was a multicenter, retrospective, observational study. Children aged 28 days to 18 years scheduled for anesthesia services at 12 centers requiring universal COVID-19 testing from March 29, 2020 to June 30, 2020 were included. COVID-19 positivity rates among those tested were plotted and trends were assessed using the Cochran Armitage test of trend. Independent risk factors were explored using multivariable logistic regression.Data were collected and analyzed on 33,320 anesthesia encounters including 265 children with COVID-19. Over the study period, the rates of infections in the pediatric anesthesia population did not demonstrate a significant trend. In the general population, there was a significant downward trend in infection rates (P.001). In exploratory analysis, multivariable risk factors for a COVID-19 positive test were Black/African American race, Hispanic ethnicity, American Society of Anesthesiologists (ASA) physical status III or above, overweight and obese body mass index (BMI), orthopedic cases, abdominal cases, emergency cases, absence of injury and trauma, and West region (all P.05).Rates of COVID-19 in pediatric anesthesia patients were consistently lower than in the general population. Independent risk factors of a positive test for children were identified. This is the first time universal testing for a single infectious disease was undertaken on a wide scale. As such, the association of infection with surgical case type or emergency case status is unprecedented.

Point-of-care hemoglobin testing devices play an important role in intraoperative anesthetic management where significant hemorrhage is anticipated; however, the reliability of these devices has not been examined in the context of pediatric liver transplantation. In this retrospective observational study, we aimed to determine whether 95% of results from two point-of-care hemoglobinometers, the HemoCue and iSTAT, would fall within a difference of ±1 g/dl, our a priori-defined clinically acceptable level of agreement, of the hemoglobin measures on a core laboratory complete blood count.We retrospectively collected data from 70 patients presenting for a liver transplant at a single center, tertiary care pediatric hospital over a 3.5-year period. We analyzed 92 contemporaneous pairs of hemoglobin values from the HemoCue and complete blood count, and 252 pairs of hemoglobin values from the iSTAT and complete blood count. Agreement between the point-of-care devices and complete blood count was assessed using Bland-Altman analysis, which was the primary outcome. Secondary analyses included an error grid analysis and Cohen's kappa statistic.Both point-of-care devices underestimated complete blood count hemoglobin values and neither device satisfied our a priori-defined clinically acceptable level of agreement that 95% of values would fall within ±1 g/dl of the complete blood count measurement. The mean difference [limits of agreement] of the HemoCue was 0.4 g/dl (p .001) [-0.9 to 1.6 g/dl] and of the iSTAT was 0.6 g/dl (p .001) [-1.4 to 2.6 g/dl]. Secondary error grid analysis revealed that neither device performed well enough to replace a complete blood count at critical thresholds of hemoglobin values.While the HemoCue and iSTAT contribute information in a timely manner during dynamic intraoperative situations, there is significant imprecision compared to the gold standard complete blood count. If clinical stability allows, the results of these point-of-care hemoglobinometers should be confirmed with a complete blood count, rather than being used as the sole factor in determining transfusion needs during pediatric liver transplantation.

In 2012, the American Academy of Pediatrics (AAP) concluded the health benefits of circumcision during the neonatal period outweigh the risks. This study describes recent trends in male circumcision in freestanding children's hospitals in the United States.Using the Pediatric Health Information System (PHIS), male patients18 years of age who were circumcised without any additional procedures between the years 2010 and 2017 were identified. Data included age at procedure (neonate: 0-30 days, infant: 31-365 days, early childhood: ≥1 to5 years, and older child: ≥5 to18 years), cost, and specialty performing the circumcision.Of the 171,680 circumcisions performed, 85,270 (50%) were during neonatal period, 29,060 (17%) during infancy, 30,276 (18%) early childhood, and 26,355 (16%) thereafter. Circumcision in neonates increased from 39% to 58% (p 0.001), and the proportion performed during infancy decreased over time. System level cost for ambulatory circumcision averaged $32 million USD annually, and median cost per ambulatory circumcision was $2892 USD. Obstetricians and Pediatricians are performing proportionally more circumcisions.Since 2012, proportionally more neonates are undergoing circumcision in US children's hospitals. Perinatal specialties are performing an increasing proportion of circumcisions. Circumcision during the birth hospitalization in the neonatal period is more resource-effective than postponing until later in infancy.Retrospective, cross-sectional analysis.Level IV.

Though there is a well-established gender pay gap in medicine, studies on compensation disparities between women and men chief executive officers (CEO) showed mixed results. We conducted a cross-sectional study of children’s hospitals in the USA to evaluate whether CEO gender was associated with compensation differences. Nine out of 31 children’s hospitals employed a female CEO. There was no significant difference between men and women CEOs in terms of hospital characteristics (location, size or ranking in US News and World report) or CEO characteristics (advanced degrees, tenure or compensation). Gender was not associated with significant differences in CEO compensation.

Background Preoperative hyponatraemia is an independent risk factor for postoperative mortality in adults. To our knowledge, this has not been investigated in children. Methods Using data from the 2014 and 2015 data sets of the American College of Surgeons National Surgical Quality Improvement Program-Pediatric (NSQIP-P), we conducted a retrospective study of children undergoing surgery. The primary outcome was 30-day all-cause mortality. The secondary outcomes of interest were postoperative seizure within 30 days and prolonged length of stay. To identify any independent association between preoperative hyponatraemia, defined as mild (serum sodium of 131–135 mEq L−1) or severe (≤130 mEq L−1), and death, postoperative seizures, or prolonged length of stay, multivariable logistic regression models were generated. Results A total of 152 894 patients were identified, and of these 35 291 were included in the final analysis. Preoperative hyponatraemia was present in 5422 patients or 15.4% of the final cohort. There were 432 (0.80%) deaths at 30 days. Compared with patients with a normal preoperative sodium concentration, those with mild (P=0.003; odds ratio [OR]: 1.59; 95% confidence interval [CI]: 1.17–2.18) and severe (P=0.002; OR: 2.16; 95% CI: 1.32–3.54) hyponatraemia had increased rates of death, after adjusting for co-morbidity and procedural complexity. Both mild (P Conclusions This retrospective analysis identified an association between preoperative hyponatraemia and perioperative mortality and length of stay in paediatric patients.

BackgroundTo date, the independent predictors and outcomes of unplanned postoperative intubation (UPI) in pediatric patients after noncardiac surgery are yet to be characterized. The authors aimed to identify the incidence and predictors of this event and evaluated the effect of this event on postoperative mortality.MethodsData of 87,920 patients from the American College of Surgeons National Surgical Quality Improvement Program Pediatric database were analyzed and assigned to derivation (n = 58,614; 66.7%) or validation (n = 29,306; 33.3%) cohorts. The derivation cohort was analyzed for the incidence and independent predictors of early UPI. The final multivariable logistic regression model was validated using the validation cohort.ResultsEarly UPI occurred with an incidence of 0.2% in both cohorts. Among the 540 patients who experienced a UPI, 178 (33.0%) were intubated within the first 72 h after surgery. The final logistic regression model indicated operation time, severe cardiac risk factors, American Society of Anesthesiologists physical status classification more than or equal to 2, tumor involving the central nervous system, developmental delay/impaired cognitive function, past or current malignancy, and neonate status as independent predictors of early UPI. Having an early UPI was associated with an increased risk of unadjusted, all-cause 30-day mortality, demonstrating an odds ratio of 11.4 (95% CI, 5.8 to 22.4).ConclusionsPediatric patients who experienced an early UPI after noncardiac surgery had an increased likelihood of unadjusted 30-day mortality by more than 11-fold. Identification of high-risk patients can allow for targeted intervention and potential prevention of such outcomes.

Background There is a paucity of data regarding risk stratification of pediatric patients presenting for low-risk skin and soft tissue surgery. Aims We sought to determine the incidence and independent predictors of postoperative complications and unplanned 30-day readmission in a cohort of children undergoing low-risk skin and soft tissue surgery. Methods The study included pediatric patients who underwent minor procedures of the skin and soft tissue at continuously enrolled American College of Surgeons National Surgical Quality Improvement Program Pediatric hospitals over a two-year period. The primary outcome was a 30-day postoperative complication composite. The secondary outcome was unplanned 30-day readmission. Results The final analysis included 6,730 patients. There were a total of 170 postoperative complications among 152 patients (2.23%) with the majority of complications being either wound-related or postoperative mechanical ventilation. The independent predictors for an increased risk of postoperative complication were American Society of Anesthesiologists classification ≥3 and nutritional deficiency. There were 41 unplanned readmissions (0.61%). The presence of a postoperative wound complication or a postoperative pulmonary complication during the index hospital stay was an independent risk factor for unplanned 30-day readmission. Conclusion Pediatric patients with American Society of Anesthesiologists classification ≥3 and nutritional deficiency undergoing low-risk surgery are at risk for the development of postoperative complications. Patients who develop wound and postoperative pulmonary complications are at higher risk for unplanned 30-day readmission. Identification of these higher risk patients may allow the anesthesiologist to implement targeted therapies to minimize the likelihood of occurrence of these complications.

Neuraxial anesthesia has been demonstrated to be safe and effective for children undergoing subumbilical surgery. There is limited evidence regarding the safety of neuraxial anesthesia in pediatric patients with a ventriculoperitoneal shunt. We evaluated a series of 25 patients with indwelling ventriculoperitoneal shunts for complications within 30 days of any procedure performed with a neuraxial technique. One patient required a ventriculoperitoneal shunt revision 5 days after a lumbar catheter placement. The neurosurgeon determined the revision to be likely unrelated to the patient's lumbar catheter. Concerns about the use of neuraxial anesthesia in patients with an indwelling ventriculoperitoneal shunt may be overstated.

To date, the independent predictors and outcomes of unplanned postoperative intubation (UPI) in pediatric patients after noncardiac surgery are yet to be characterized. The authors aimed to identify the incidence and predictors of this event and evaluated the effect of this event on postoperative mortality.Data of 87,920 patients from the American College of Surgeons National Surgical Quality Improvement Program Pediatric database were analyzed and assigned to derivation (n = 58,614; 66.7%) or validation (n = 29,306; 33.3%) cohorts. The derivation cohort was analyzed for the incidence and independent predictors of early UPI. The final multivariable logistic regression model was validated using the validation cohort.Early UPI occurred with an incidence of 0.2% in both cohorts. Among the 540 patients who experienced a UPI, 178 (33.0%) were intubated within the first 72 h after surgery. The final logistic regression model indicated operation time, severe cardiac risk factors, American Society of Anesthesiologists physical status classification more than or equal to 2, tumor involving the central nervous system, developmental delay/impaired cognitive function, past or current malignancy, and neonate status as independent predictors of early UPI. Having an early UPI was associated with an increased risk of unadjusted, all-cause 30-day mortality, demonstrating an odds ratio of 11.4 (95% CI, 5.8 to 22.4).Pediatric patients who experienced an early UPI after noncardiac surgery had an increased likelihood of unadjusted 30-day mortality by more than 11-fold. Identification of high-risk patients can allow for targeted intervention and potential prevention of such outcomes.

The role of 5-lipoxygenase (5-LO) in colon cancer is unknown. Tumor-infiltrating macrophages, neutrophils, and mast cells have been shown to play important roles in colon tumorigenesis and are dependent on 5-LO for function.Utilizing the APC(Δ468) polyposis model, we performed 5-LO gene knockouts and evaluated the subsequent changes in macrophage, neutrophil, and mast cell density at the tumor site. The proliferative and degranulation capacities of 5-LO-deficient mast cells were also measured, quantifying thymidine incorporation and β-hexosaminidase release, respectively.APC(Δ468)/5LO(-/-) mice displayed increased tumor-infiltrating macrophages and decreased neutrophils at the polyp site. In vitro, mast cells deficient for 5-LO proliferated at a diminished rate while mast cell degranulation was unchanged.We provide evidence suggesting that 5-LO deficiency has differential effects on the infiltration of macrophages and neutrophils in adenomatous polyps, increasing and decreasing infiltration of these cells, respectively. Our observations are consistent with a protective role for tumor-infiltrating macrophages in the initiation of polyp formation. The mechanisms through which 5-LO deficiency negatively affects these cells are under investigation.These results provide evidence that 5-LO plays an important role in tumorigenesis and further indicate that 5-LO-selective inhibitors can be investigated as potential therapeutic agents for colorectal polyposis and cancer.

Pancreatic cancer is associated with a pronounced fibrotic reaction that was recently shown to limit delivery of chemotherapy. To identify potential therapeutic targets to overcome this fibrosis, we examined the interplay between fibrosis and the key proteinase membrane type 1-matrix metalloproteinase (MT1-MMP, MMP-14), which is required for growth and invasion in the collagen-rich microenvironment. In this article, we show that compared with control mice (Kras+/MT1-MMP−) that express an activating KrasG12D mutation necessary for pancreatic cancer development, littermate mice that express both MT1-MMP and KrasG12D (Kras+/MT1-MMP+) developed a greater number of large, dysplastic mucin-containing papillary lesions. These lesions were associated with a significant amount of surrounding fibrosis, increased α-smooth muscle actin (+) cells in the stroma, indicative of activated myofibroblasts, and increased Smad2 phosphorylation. To further understand how MT1-MMP promotes fibrosis, we established an in vitro model to examine the effect of expressing MT1-MMP in pancreatic ductal adenocarcinoma (PDAC) cells on stellate cell collagen deposition. Conditioned media from MT1-MMP–expressing PDAC cells grown in three-dimensional collagen enhanced Smad2 nuclear translocation, promoted Smad2 phosphorylation, and increased collagen production by stellate cells. Inhibiting the activity or expression of the TGF-β type I receptor in stellate cells attenuated MT1-MMP conditioned medium–induced collagen expression by stellate cells. In addition, a function-blocking anti–TGF-β antibody also inhibited MT1-MMP conditioned medium–induced collagen expression in stellate cells. Overall, we show that the bona fide collagenase MT1-MMP paradoxically contributes to fibrosis by increasing TGF-β signaling and that targeting MT1-MMP may thus help to mitigate fibrosis. Mol Cancer Res; 9(10); 1294–304. ©2011 AACR.

Bevacizumab (Avastin™; rhuMab VEGF), a monoclonal antibody targeting vascular endothelial growth factor (VEGF), has seen increased use in the perioperative treatment of colorectal and pancreatic cancer. Little is known, however, regarding its impact on surgical outcomes in patients undergoing resection. The objective of this review was to examine if the addition of bevacizumab to existing neoadjuvant regimens increases morbidity after cancer resection.

T-regulatory cells (Treg) and mast cells (MC) are abundant in colorectal cancer (CRC) tumors. Interaction between the two is known to promote immune suppression or loss of Treg functions and autoimmunity. Here, we demonstrate that in both human CRC and murine polyposis the outcome of this interaction is the generation of potently immune suppressive but proinflammatory Treg (ΔTreg). These Treg shut down IL10, gain potential to express IL17, and switch from suppressing to promoting MC expansion and degranulation. This change is also brought about by direct coculture of MC and Treg, or culture of Treg in medium containing IL6 and IL2. IL6 deficiency in the bone marrow of mice susceptible to polyposis eliminated IL17 production by the polyp infiltrating Treg, but did not significantly affect the growth of polyps or the generation of proinflammatory Treg. IL6-deficient MC could generate proinflammatory Treg. Thus, MC induce Treg to switch function and escalate inflammation in CRC without losing T-cell–suppressive properties. IL6 and IL17 are not needed in this process.

Purpose Leukotrienes and prostaglandins, products of arachidonic acid metabolism, sustain both systemic and lesion-localized inflammation. Tumor-associated Inflammation can also contribute to the pathogenesis of colon cancer. Patients with inflammatory bowel disease (IBD) have increased risk of developing colon cancer. The levels of 5-lipoxygenase (5-LO), the key enzyme for leukotrienes production, are increased in colon cancer specimens and colonic dysplastic lesions. Here we report that Zileuton, a specific 5-LO inhibitor, can prevent polyp formation by efficiently reducing the tumor-associated and systemic inflammation in APCΔ468 mice. Experimental Design In the current study, we inhibited 5-LO by dietary administration of Zileuton in the APCΔ468 mouse model of polyposis and analyzed the effect of in vivo 5-LO inhibition on tumor-associated and systemic inflammation. Results Zileuton-fed mice developed fewer polyps and displayed marked reduction in systemic and polyp-associated inflammation. Pro-inflammatory cytokines and pro-inflammatory innate and adaptive immunity cells were reduced both in the lesions and systemically. As part of tumor-associated inflammation Leukotriene B4 (LTB4), product of 5-LO activity, is increased focally in human dysplastic lesions. The 5-LO enzymatic activity was reduced in the serum of Zileuton treated polyposis mice. Conclusions This study demonstrates that dietary administration of 5-LO specific inhibitor in the polyposis mouse model decreases polyp burden, and suggests that Zileuton may be a potential chemo-preventive agent in patients that are high-risk of developing colon cancer.

Mast cells (MC) are a bone marrow-derived, long-lived, heterogeneous cellular population that function both as positive and negative regulators of immune responses. They are arguably the most productive chemical factory in the body and influence other cells through both soluble mediators and cell-to-cell interaction. MC are commonly seen in various tumors and have been attributed alternatively with tumor rejection or tumor promotion. Tumor-infiltrating MC are derived both from sentinel and recruited progenitor cells. MC can directly influence tumor cell proliferation and invasion but also help tumors indirectly by organizing its microenvironment and modulating immune responses to tumor cells. Best known for orchestrating inflammation and angiogenesis, the role of MC in shaping adaptive immune responses has become a focus of recent investigations. MC mobilize T cells and antigen-presenting dendritic cells. They function as intermediaries in regulatory T cells (Treg)-induced tolerance but can also modify or reverse Treg-suppressive properties. The central role of MC in the control of innate and adaptive immunity endows them with the ability to tune the nature of host responses to cancer and ultimately influence the outcome of disease and fate of the cancer patient.

Arachidonic acid metabolism has been implicated in colon carcinogenesis, but the role of hematopoietic 5-lipoxygenase (5LO) that may impact tumor immunity in development of colon cancer has not been explored. Here we show that tissue-specific deletion of the 5LO gene in hematopoietic cells profoundly attenuates polyp development in the APCΔ468 murine model of colon polyposis. In vitro analyses indicated that mast cells in particular utilized 5LO to limit proliferation of intestinal epithelial cells and to mobilize myeloid-derived suppressor cells (MDSCs). Mice lacking hemapoietic expression of 5LO exhibited reduced recruitment of MDSCs to the spleen, mesenteric lymph nodes, and primary tumor site. 5LO deficiency also reduced the activity in MDSCs of arginase-1, which is thought to be critical for MDSC function. Together, our results establish a pro-tumorigenic role of hematopoietic 5LO in the immune microenvironment and suggest 5LO inhibition as an avenue for future investigation in treatment of colorectal polyposis and cancer. Cancer Res; 71(5); 1627–36. ©2011 AACR.

Diets containing omega-6 (ω-6) fat have been associated with increased tumor development in carcinogen-induced pancreatic cancer models. However, the effects of ω-6 fatty acids and background strain on the development of genetically-induced pancreatic neoplasia is unknown. We assessed the effects of a diet rich in ω-6 fat on the development of pancreatic neoplasia in elastase (EL)-Kras(G12D) (EL-Kras) mice in two different backgrounds. EL-Kras FVB mice were crossed to C57BL/6 (B6) mice to produce EL-Kras FVB6 F1 (or EL-Kras F1) and EL-Kras B6 congenic mice. Age-matched EL-Kras mice from each strain were compared to one another on a standard chow. Two cohorts of EL-Kras FVB and EL-Kras F1 mice were fed a 23% corn oil diet and compared to age-matched mice fed a standard chow. Pancreata were scored for incidence, frequency, and size of neoplastic lesions, and stained for the presence of mast cells to evaluate changes in the inflammatory milieu secondary to a high fat diet. EL-Kras F1 mice had increased incidence, frequency, and size of pancreatic neoplasia compared to EL-Kras FVB mice. The frequency and size of neoplastic lesions and the weight and pancreatic mast cell densities in EL-Kras F1 mice were increased in mice fed a high ω-6 fatty acid diet compared to mice fed a standard chow. We herein introduce the EL-Kras B6 mouse model which presents with increased frequency of pancreatic neoplasia compared to EL-Kras F1 mice. The phenotype in EL-Kras F1 and FVB mice is promoted by a diet rich in ω-6 fatty acid.

Purpose: To assess the clinical and pathologic significance of mast cell infiltration in human pancreatic cancer and evaluate crosstalk between mast cells and cancer cells in vitro. Experimental Design: Immunohistochemistry for tryptase was done on 53 pancreatic cancer specimens. Mast cell counts were correlated with clinical variables and survival. Serum tryptase activity from patients with cancer was compared with patients with benign pancreatic disease. In vitro, the effect of pancreatic cancer–conditioned medium on mast cell migration was assessed. The effect of conditioned medium from the human mast cell line, LAD-2, on cancer and normal ductal cell proliferation was assessed by thymidine incorporation. Matrigel invasion assays were used to evaluate the effect of mast cell–conditioned medium on cancer cell invasion in the presence and absence of a matrix metalloproteinase inhibitor, GM6001. Results: Mast cell infiltration was significantly increased in pancreatic cancer compared with normal pancreatic tissue (11.4 ± 6.7 versus 2.0 ± 1.4, P < 0.001). Increased infiltrating mast cells correlated with higher grade tumors (P < 0.0001) and worse survival. Patients with pancreatic cancer had elevated serum tryptase activity (P < 0.05). In vitro, AsPC1 and PANC-1 cells induced mast cell migration. Mast cell–conditioned medium induced pancreatic cancer cell migration, proliferation, and invasion but had no effect on normal ductal cells. Furthermore, the effect of mast cells on cancer cell invasion was, in large part, matrix metalloproteinase–dependent. Conclusions: Tumor-infiltrating mast cells are associated with worse prognosis in pancreatic cancer. In vitro, the interaction between mast cells and pancreatic cancer cells promotes tumor growth and invasion. Clin Cancer Res; 16(8); 2257–65. ©2010 AACR.

To dissect the role of constitutively altered Tgfbr1 signaling in pancreatic cancer development, we crossed Elastase-KrasG12D (EL-Kras) mice with Tgfbr1 haploinsufficient mice to generate EL-Kras/Tgfbr1+/− mice. Mice were euthanized at 6 to 9 months to compare the incidence, frequency, and size of precancerous lesions in the pancreas. Only 50% of all EL-Kras/Tgfbr1+/− mice developed preinvasive lesions compared with 100% of EL-Kras (wild-type Tgfbr1) mice. The frequency of precancerous lesions was 4-fold lower in haploinsufficient than in control mice. Paradoxically, the precancerous lesions of EL-Kras/Tgfbr1+/− mice were considerably larger than those in EL-Kras mice. Yet, the mitotic index of precancerous cells and the observable levels of fibrosis, lipoatrophy, and lymphocytic infiltration were reduced in EL-Kras/Tgfbr1+/− mice. We conclude that Tgfbr1 signaling promotes the development of precancerous lesions in mice. These findings suggest that individuals with constitutively decreased TGFBR1 expression may have a decreased risk of pancreatic cancer. [Cancer Res 2009;69(24):9169–74]